Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:5.99.1.2 (topoisomerase)
 9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A number of characteristics in the human genetic disorder ataxia-telangiectasia are compatible with an alteration to chromatin structure or the recognition of that structure by an enzyme or DNA binding protein. We describe here reduce activity of DNA topoisomerase type II in a number of Epstein Barr Virus-transformed ataxia-telangiectasia lymphoblastoid cell lines. Enzyme activity was reduced 10-fold or greater in 4 out of 5 cell lines compared to controls. In the remaining cell line approximately a 2-3 fold reduction was evident in partially purified extracts. DNA topoisomerase type I activity was found to be the same as controls in all the cell lines. Northern blot analysis revealed that the same level of DNA topoisomerase II mRNA was expressed in ataxia-telangiectasia and control cell lines. The size and amount of the enzyme did not differ appreciably from that observed in control cells. The reduced activity of DNA topoisomerase II in ataxia-telangiectasis cells might be explained by amino acid substitutions, small deletions in DNA or by a defect in post-translational modification in these cells.
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PMID:Defective DNA topoisomerase II in ataxia-telangiectasia cells. 196 59

We have provided evidence recently for a defect in DNA topoisomerase II in ataxia--telangiectasia (A-T) lymphoblastoid cells. This study was initiated to investigate in greater detail the nature of this defect. Southern hybridization analysis was carried out on DNA from control and A-T Epstein--Barr virus-transformed lymphoblastoid cells. The pattern of digestion, using several restriction enzymes, was the same in both cell types. Expression of topoisomerase II mRNA occurred to the same extent and there was no difference in the size of mRNA between the cell types. Western blot analysis revealed that the same amount of a major band of topoisomerase II protein was present in A-T and control cells but there was evidence for a reduced amount of a lower-molecular-weight form in A-T only. Extraction and purification did not lead to alteration in size of the enzyme or in amount recovered.
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PMID:Study of DNA topoisomerase II in ataxia-telangiectasia cells. 247 31

Abnormal expression of the nuclear-associated enzyme DNA topoisomerase II (topoisomerase II) has been implicated in the in vitro phenotype of radiation hypersensitive ataxia-telangiectasia (A-T) cells and in modifying sensitivity of eukaryotic cells to topoisomerase II-inhibitor drugs [e.g., the DNA intercalator amsacrine (mAMSA)]. To study such relationships, various SV40- and Epstein-Barr Virus-transformed human cell lines derived from normal, A-T, or UV-sensitive xeroderma pigmentosum donors have been assayed for their sensitivity to mAMSA together with direct and indirect measurements of topoisomerase II expression. We report on the identification of an SV40-transformed A-T fibroblast cell line with abnormally high levels of topoisomerase II in nuclear protein extracts as determined by immunoblotting, measurement of kinetoplast DNA decatenation activity, and mAMSA-dependent DNA-protein cross-linking activity in a filter binding assay. Using a flow cytometric assay for the analysis of reactivity of nuclei with a polyclonal antitopoisomerase II antibody, overproduction was found to occur in all phases of the cell cycle. High levels of topoisomerase II were associated with hypersensitivity (5-10-fold) to mAMSA-induced cell cycle delay, cell kill, and DNA strand breakage (assayed under protein-denaturing conditions). Xeroderma pigmentosum (group A) cells demonstrated normal responses to mAMSA. The results provide evidence that cellular potential for the generation of topoisomerase II-dependent DNA damage is a major factor in governing the sensitivity to mAMSA. Furthermore, underexpression of topoisomerase II does not appear to be a primary factor in describing the in vitro A-T phenotype. The findings also relate to how changes in chromatin structure and function may either reflect or dictate the expression of topoisomerase II in human cells.
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PMID:Cellular consequences of overproduction of DNA topoisomerase II in an ataxia-telangiectasia cell line. 253 42

Considerable evidence supports a defect at the level of chromatin structure or recognition of that structure in cells from patients with the human genetic disorder ataxia-telangiectasia. Accordingly, we have investigated the activities of enzymes that alter the topology of DNA in Epstein Barr Virus-transformed lymphoblastoid cells from patients with this syndrome. Reduced activity of DNA topoisomerase II, determined by unknotting of P4 phage DNA, was observed in partially purified extracts from 5 ataxia-telangiectasia cell lines. The levels of enzyme activity was reduced substantially in 4 of these cell lines and to a lesser extent in the other cell line compared to controls. DNA topoisomerase I, assayed by relaxation of supercoiled DNA, was found to be present at comparable levels in both cell types. Reduced activity of topoisomerase II in ataxia-telangiectasia is compatible with the molecular, cellular and clinical changes described in this syndrome.
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PMID:Reduced DNA topoisomerase II activity in ataxia-telangiectasia cells. 283 4

The efficiency of stable transformation of human cells by integrative (non-replicating) plasmids carrying a selectable gene has been shown to be markedly enhanced by the introduction into the plasmid DNA of bulky damage, such as cyclobutane pyrimidine dimers or psoralen photoadducts. Enhanced transformation (ET) occurs in all human cells tested, including DNA repair-deficient cells from the hereditary syndrome xeroderma pigmentosum, but significantly less, if at all, in rodent cells. ET has been observed with a variety of integrative plasmid constructs, suggesting the generality of the phenomenon; as expected, ET is due to an increase in the number of cells carrying integrated plasmid sequences. In contrast to integrative plasmids, stable transformation by episomal (autonomously replicating) plasmids derived from the Epstein-Barr virus is only depressed by the introduction of photoproducts; furthermore, pronounced inactivation of transformation mediated by episomal plasmids becomes apparent in xeroderma pigmentosum cells. Altogether, these results suggest that DNA damage increases the probability of stable insertion of heterologous non-replicating DNA into human chromosomes. Moreover, the differential sensitivity to DNA damage of human cell transformation mediated by integrative versus episomal plasmids suggests caution in using such assay to measure host cell reactivation capacity; processing of DNA damage in mammalian cells might differ significantly between intra- versus extra-chromosomal DNA. Since ET may be induced by damage outside the selectable gene carried on integrative plasmids, we propose a model that involves local disruption of chromatin structure by helix-distorting DNA lesions flanking actively transcribed sequences; alternatively, reorganization of such altered DNA structure might be favored by the presence of topoisomerase-like activities in the proximity of active genes. Because ET can also be induced by DNA damage to the recipient cells, it is speculated that similar mechanism(s) might be involved in the generation of other types of non-homologous DNA recombination in damaged human chromosomes, including oncogenic cell transformation mediated by integrative DNA viruses.
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PMID:Effect of DNA damage on stable transformation of mammalian cells with integrative and episomal plasmids. 292 24

The studies described below were carried out to analyze the damage induced by DNA active drugs to episomal (Epstein-Barr virus, EBV) DNA in the Raji Burkitt's lymphoma cell line. This work: (i) applies pulsed-field gel electrophoresis (PFGE) techniques to quantify DNA damage on a large (approximately 180 kbp), circular target, (ii) investigates the DNA strand-scission behavior of different classes of drugs on the EBV episome, and (iii) compares EBV episomal damage to that generated in genomic DNA in the Raji cell line. Cells were treated with ionizing radiation to induce random strand scission, and the migration of topological forms of EBV was measured using PFGE. DNA damage induced in the episome by DNA active drugs was then assayed. Three drugs, acting by different types of DNA interactive mechanisms, were used: bleomycin, an intercalative DNA strand-scission agent; and amsacrine (mAMSA) and teniposide (VM26), intercalative and nonintercalative topoisomerase II active drugs, respectively. Rad equivalency of damage was determined by comparing the drug-induced change in percentage of Forms I and III to that generated by ionizing radiation. Additionally, single- and double-strand scission induced in genomic (total cellular) DNA by X-rays, bleomycin, amsacrine, and teniposide were assayed by high-sensitivity alkaline and neutral filter elution techniques. We demonstrate that pulsed-field gel electrophoresis is a useful technique for measuring form conversion in large episomal DNA. While all three drugs effect both episomal and genomic DNA strand scission, bleomycin appears to preferentially damage the EBV episome. The topoisomerase II active drugs mAMSA and VM26 show no evidence of episome-directed damage in this system and, in fact, damage genomic DNA at somewhat higher rates.
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PMID:Damage induced in episomal EBV DNA in Raji cells by antitumor drugs as measured by pulsed field gel electrophoresis. 752 29

The roles of topoisomerases I and II in Epstein-Barr virus (EBV) replication were investigated using Raji cells infected with EBV. The topoisomerase II inhibitor ellipticine inhibited the synthesis of EBV polypeptides at concentrations which did not affect total protein synthesis. Slot blot analysis of total cellular DNA showed that camptothecin and ellipticine inhibited replication of progeny EBV DNA in superinfected Raji cells at concentrations which did not inhibit synthesis of EBV early polypeptides prerequisite for EBV DNA replication. Analysis of the structure of EBV DNA termini demonstrated that both inhibitors affected the replicating EBV DNA. Gardella gel electrophoresis showed that both inhibitors affected the formation of the linear form of EBV DNA. However, restriction analysis of EBV DNA in superinfected Raji cells demonstrated that both inhibitors degraded neither endogenous nor exogenous EBV DNA. Cell viability was not affected by either inhibitor at the concentrations tested. These findings suggest that topoisomerase II is required for expression of the EBV genome and that both topoisomerases I and II are involved in replication of the EBV genome during the lytic phase of the life cycle. The effects of topoisomerase inhibitors on the circular form of EBV DNA during virus replication are discussed.
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PMID:Topoisomerase I and II activities are required for Epstein-Barr virus replication. 840 49

We studied the effects of nine cytotoxic drugs on three groups of B-lymphoblastoid cell lines transformed by Epstein-Barr virus (EBV): group 1, mortal cell lines from normal individuals; group 2, immortalized cell lines from normal individuals with strong telomerase activity; group 3, mortal cell lines from Werner's syndrome (WS) patients. Aminoglycoside antibiotics and alkylating drugs showed significantly stronger cytotoxic effects on immortalized cell lines than on mortal cell lines or the cell lines before immortalization. In contrast, topoisomerase II inhibitors showed no difference or they tended to be less cytotoxic to immortalized cell lines. Mortal cell lines from normal individuals and WS patients showed no difference in sensitivity against all the drugs examined except for the topoisomerase I inhibitor, camptothecin, which had a stronger cytotoxic effect on WS cell lines than other cell lines. We discuss the mechanisms underlying these cytotoxic effects.
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PMID:Differential effects of cytotoxic drugs on mortal and immortalized B-lymphoblastoid cell lines from normal and Werner's syndrome patients. 955 52

Epstein-Barr virus (EBV) is associated with a number of human malignancies. In vitro EBV infection transforms human lymphocytes into proliferating cell lines (EBV-lymphocytes). Etoposide, topoisomerase II inhibitor, induced apoptosis in EBV-lymphocytes as shown by expression of phosphatidylserine, loss of DNA and mitochondrial membrane potential, and cell shrinkage. In contrast, those cells, which had yet to display signs of apoptosis, grew to exceed their normal size. These EBV-lymphocytes had unusual cellular and nuclear morphology, higher mitochondrial membrane potential, increased expression of proteins and an amount of DNA that exceeded the maximum DNA content in normal EBV-lymphocytes by more than two-fold. Application of the caspase inhibitor Z-VAD-FMK in the presence of etoposide increased the numbers of such large cells. This data suggests that inhibition of topoisomerase II by etoposide does not inhibit DNA synthesis but rather overrides the G(2)/M check points of the cell cycle, resulting in cells growth over their genetically determined size. This may trigger apoptosis to eliminate cells, which failed to complete mitosis.
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PMID:Inhibition of topoisomerase II overrides the G2/M check points of the cell cycle in EBV-lymphocytes. 1281 83

We have shown previously that the Epstein-Barr virus nuclear antigen-1 (EBNA1) can act as a transforming suppressor in the HER2/neu-overexpressing ovarian cancer cells. In the present study, by using flow cytometric analysis, we demonstrate that EBNA1 could prolong G(2)/M phase and sensitize to Taxol-induced apoptosis in the EBNA1-expressing ovarian cancer cell stable transfectants. In addition, EBNA1 could also significantly increase topoisomerase IIalpha protein expression, indicating that the up-regulation of topoisomerase IIalpha may be one of the mechanisms by which EBNA1 enhances the sensitivity of ovarian cancer cells to topoisomerase II-targeting anticancer drugs, such as VP-16 and Adriamycin. These data suggest that EBNA1 not only prolongs cell cycle at G(2)/M phase and up-regulates topoisomerase IIalpha expression in HER2/neu-overexpressing ovarian cancer cells, but also increases cellular apoptosis through sensitization of cancer cells to topoisomerase II-directing anticancer drugs.
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PMID:EBNA1 may prolong G(2)/M phase and sensitize HER2/neu-overexpressing ovarian cancer cells to both topoisomerase II-targeting and paclitaxel drugs. 1289 73


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